The rod cells of vertebrate retinas are responsible for vision at low levels of light. Rod outer segment disks are stacked along the length of the outer segment. The disks are formed from evaginations of the plasma membrane at the base of the outer segment. They are displaced toward the apical Lip as new disks are formed. The disk membranes can now be separated as a function of their cholesterol content using sucrose density gradients. High levels of membrane cholesterol are associated with newly synthesized disks. Both the phospholipid headgroup and the total fatty acid composition remains essentially constant among the disks as they age. Thus the major change observed in the lipid composition of disk membranes as they are apically displaced is a decrease in the membrane cholesterol content. However new disks and plasma membrane differ significantly in phospholipid composition. This suggests that an important sorting process occurs at disk formation. By completing the lipid and protein analysis of plasma membrane and of disks as a function of age we will obtain some understanding of this interesting sorting process. It is possible that these sorting processes are impaired or altered in disease states. Therefore these studies will be extended to other animals, including a comparison of normal and dystrophic rats. The visual pigment, rhodopsin, is an integral membrane protein of the disks and functions in the primary event of visual transduction. Rhodopsin initiates a cascade of events which culminates in the hydrolysis of cGMP and the closure of the plasma membrane Na+ channels. The membrane cholesterol composition is capable of modulating this membrane activity. In particular the loss of cholesterol from the disk membrane would appear, from preliminary data to increase the potential response to light. Therefore the dependence of membrane activity on cholesterol will be examined in greater detail during this project period. The effect of cholesterol on rhodopsin properties such as photolysis, regeneration and thermal stability will be investigated. It is anticipated that this information will provide insight with respect to the mechanism of the cholesterol effect. Finally, the possibility that cholesterol may be responsible, at least in part, for the significantly reduced sensitivity of some cones cells, relative to rod cells, will be investigated. Cone cells in which the "disks" are not separated from the plasma membrane do not have the opportunity to move the photopigments to a reduced cholesterol environment with its apparently increased sensitivity to light.